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Phire® Animal Tissue Direct PCR Kit
Recommended sample sizes
The sample size recommendations below are based on organisms and tissue types that have been tested so far. Some optimization may be needed when starting to perform Direct PCR reactions on a new sample, but the recommendations can be used as guidelines.
In general, it is beneficial to take as small a piece of the sample as possible to minimize the amount of inhibitors in the reaction.
Note: Direct protocol is recommended only for short amplicons (< 500 bp for fish fins and < 1kb for other tissues).
| Tissue sample | Direct protocol (50 µl reaction) |
Dilution protocol1 (20 µl Dilution Buffer + 0.5 µl DNARelease Additive) |
|
|---|---|---|---|
| C. elegans | 2-25 worms | 25-40 worms2 | |
| Drosophila | Wing (e.g. 1 mm x 2 mm) | Whole Drosophila | |
| Mouse | ear | 0.5 mm punch | 2 mm punch |
| tail | 0.5 mm punch | 1-3 mm tail section | |
| embryo | - | 0.5 x 1 mm piece | |
| organs (such as spleen)3 | 1 x 1 mm piece | 1 x 1 mm piece | |
| cultured cells (=fibroblasts)4 |
- | 104 cells | |
| Zebrafish | fins (tail) | 0.5 mm punch | 1 mm x 5 mm |
| Whitefish | fins (tail) | 0.5 mm punch | 1 mm x 5 mm |
| scale | 0.5 mm punch | 1 scale (approx. 2 mm diameter) | |
| Agrilus viridis | - | One leg or 0.5 x 1 mm piece of body | |
| Animal hair | 1-2 mm region with follicle | 2-3 mm region with follicle | |
| Bird feather | 1-2 mm piece of quill end | 2-3 mm piece of quill end | |
| Muscle tissue | 0.5 mm punch | 2 mm punch | |
1) Make sure the sample is covered with solution. If larger samples are used, adjust the volumes of Dilution Buffer and DNARelease Additive accordingly.
2) If less worms are used, adjust the volume of the Dilution Buffer and DNARelease Additive accordingly.
3) The direct protocol is not recommended for fat or bone tissue.
4) The cells were trypsinated, washed with PBS and collected by centrifugation.
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| Reagents | Instruments | Consumables |
